Human equilibrative nucleoside transporter 1 (hENT1) expression is a potential predictive tool for response to gemcitabine in patients with advanced cholangiocarcinoma

Abstract

Background

Cholangiocarcinoma (CC) is a rare cancer of the liver. Surgery offers the only chance for cure. When surgery is unfeasible, chemotherapy is the backbone of treatment. The combined administration of cisplatin and gemcitabine is considered standard of care. Human equilibrative nucleoside transporter 1 (hENT1) is the major transporter responsible for gemcitabine uptake into cells. hENT1 expression is associated with an increased survival for patients receiving gemcitabine after pancreatic cancer surgery, suggesting that hENT1 is predictive of response to gemcitabine.

Aim

To determine whether there is a correlation between the expression of hENT1 and disease outcome in CC.

Methods

A retrospective study on 43 patients treated at our centre with a locally advanced or metastatic CC, who received first line treatment with gemcitabine, was performed.

Results

For the whole population, median Progression Free Survival (PFS) and overall survival (OS) were 4.0 (95% Confidence Interval 2.7–5.3 months) and 10.0 months (95% CI 6.8–13.2 months), respectively. From the 26 samples available for hENT1 staining, 18 (69%) and 8 (31%) patients had high and low hENT1 immunostaining, respectively. The median PFS were 2.0 versus 6.0 months for low versus high staining respectively (p = 0.012). The median OS were 5.0 versus 11.0 months for low versus high staining, respectively (p = 0.036). On multivariate analysis, hENT1 expression was the single independent predictive factor associated with prolonged PFS (HR 0.35, p = 0.023) and OS (HR 0.41, p = 0.046).

Conclusion

In this study we show the potential of hENT1 expression as a predictor of outcome in CC treated with gemcitabine. Larger studies are necessary to confirm these promising results.

Introduction

Cholangiocarcinoma (CC) is the second most frequent primary cancer of the liver. Worldwide, CC accounts for 3% of all gastrointestinal cancers. Its incidence has been rising over the past two decades in the United States and France.1, 2 It arises from the epithelial lining of the intrahepatic or extrahepatic bile duct. The anatomic location of CC can be described as intrahepatic, perihilar or distal extrahepatic. Five year survival rates are around 10–40% for CC.³ Complete surgical resection offers the only chance for cure. However, only 10–20% of patients present with early stage disease are considered surgical candidates. Among these resected patients, recurrence rates are high, thus for the vast majority of CC patients, systemic chemotherapy is the mainstay of their treatment plan. Patients with unresectable or metastatic CC have a poor prognosis with a median overall survival (OS) of <1 year, as shown by a comprehensive pooled analysis of published clinical trials.⁴ These authors also showed that chemotherapy with gemcitabine combined with cisplatin or oxaliplatin could increase response rate and tumour control rate in CC. Very recently, a randomised controlled phase III study, the ABC-02 study⁵, has shown a clear benefit of a combination regimen over gemcitabine alone, considered by many as the standard chemotherapy. Authors show the superiority of the combination of cisplatin and gemcitabine over gemcitabine alone, in terms of Progression Free Survival (PFS) and OS, with a nearly 4 month absolute benefit in survival and a hazard ratio of death of 0.63 (p < 0.0001). This combination can thus be considered the new standard for advanced non-operable or metastatic CC.

However, chemotherapy is administered to patients without knowledge of the genetic background of the disease, which may affect drug efficacy.

Genetically determined variability of key enzymes has been shown to influence response and toxicity of cytotoxic agents including 5-fluorouracil, irinotecan and 6-mercaptopurine.⁶ Potential candidates to predict which patients are likely to respond to treatment are genes encoding proteins involved in metabolism, transport across membranes, or target of anticancer drugs.⁷ It is likely that genetic variability of key enzymes in gemcitabine transport and metabolism may therefore have an impact on treatment response and toxicity of gemcitabine. Because the biochemical targets of gemcitabine are intracellular, the mandatory first step in the production of cytotoxicity is permeation across the cell membrane. Gemcitabine and physiologic nucleosides are hydrophilic, and diffusion through the cell membrane lipid layer is slow. Therefore, efficient cellular uptake requires the presence of specialised integral membrane nucleoside transporter proteins.⁸ Two general processes of nucleoside transport have been identified: the equilibrative bi-directional facilitators (human equilibrative nucleoside transporter 1) and the concentrative sodium/nucleoside symporters (hCNT1 and 3). The major routes for transporting gemcitabine are human equilibrative nucleoside transporter (hENT1) and to lesser extent, hCNT1 and hCNT3.9, 10, 11 Authors have shown that cells lacking hENT1 are highly resistant to gemcitabine.¹² The abundance and distribution of the hENT1 protein can be evaluated using immunohistochemistry and has been assessed in a number of malignant and benign tissues.13, 14, 15, 16, 17 Very interestingly, a recent randomised phase III trial showed that hENT1 protein expression was associated with an increased overall and disease-free survival in patients operated for a pancreatic cancer who received adjuvant gemcitabine, but not in those who received 5-FU and radiotherapy after surgery, suggesting that hENT1 expression may be predictive of response to gemcitabine.¹⁸ These results have been reproduced by others.¹⁹

On the other hand, two retrospective studies showed that overexpression of hENT1 protein was significantly correlated with poor prognosis in patients with resected ampullary and gastric cancers that did not receive any chemotherapy,13, 17 suggesting that hENT1 expression could also be a prognostic marker.

On the basis of these findings, we reviewed the files of 43 patients treated at our centre with a locally advanced or metastatic CC, out of 60 patients who received first line treatment with gemcitabine alone. The aim of the study was to determine whether there was a correlation between the expression of hENT1 and disease outcomes in CC. We excluded 17 patients treated with gemcitabine for a gallbladder cancer, because their prognosis and sensitivity to chemotherapy is different.⁴ Although increasing knowledge suggests that extrahepatic (including perihilar) CC and intrahepatic CC could have different pathophysiology, they share common molecular features, as described by a recent review,²⁰ and are treated the same way. We therefore decided to analyse together both extra- and intrahepatic CC in this study.